KS is a polyclonal malignancy that occurs at high frequency in subpopulations of HIV-infected individuals as well as in some non-HIV infected populations. Attempts as identifying an infectious co-factor in KS cells have been unfruitful, epidemiologic data suggesting such a co- factor. We have determined that KS cells differ dramatically from normal endothelial cells in their responsiveness to agents that induce VCAM-1, especially in their molecular responsiveness to double stranded RNA. This is most likely due to differential activation of the double stranded RNA activated protein kinase, PKR. This is a kinase that is important in the host anti-viral response, and many viruses have evolved mechanisms to subvert the antiviral activities of PKR. PKR is an IFN-responsive gene that has recently been shown to be a tumor suppressor gene. We hypothesize that functional alterations of PKR, possibly due to a viral inhibitor in the KS cells, are important in the pathogenesis of KS. We hypothesize that clinical responses to IFN might result from higher levels of PKR protein overcoming some of the functional inhibition of PKRs tumor suppressor function. We have demonstrated that microvascular endothelial cells transfected with either a viral inhibitor of PKR function or a mutant PKR that functionally inactivates wild type PKR develop a KS-like phenotype, supporting the hypothesis that alteration of PKR function is critical in the development of KS. In this proposal, we will fully characterize PKR induction and activation in KS cells compared to normal ECs. We will examine changes in autophosphorylation, in phosphorylation of the translation factor eIF- 2alpha, and in activation of NF-kappaB in KS cells compared to normal ECs. We will then identify the macromolecular components in KS cells that are responsible for the functional differences. We will also identify genes that are differentially expressed in KS cells compared to normal and activated microvascular endothelial cells. cDNA libraries will be created from cultured KS cells and from cultured skin microvascular endothelial cells grown under identical conditions. Differential screening of cloned genes will be undertaken to identify genes expressed exclusively in the KS cells compared to normal endothelial cells and compared to cytokine activated endothelial cells. Functional assays of these genes will be done by expressing selected sequences in cultured microvascular endothelial cells to see if they generate a KS phenotype. In situ hybridization and immunohistochemistry will be done to localize the expression of these genes in KS versus uninvolved tissue. The relationship of these genes to altered PKR activity will also be explored.